Coupling of AFM and inverted optical microscopy
Inverted optical microscopy coupled with AFM imaging offers the unique opportunity for optical data (e.g., relative ratios or fluorescence data) to be combined with high-resolution morphological images and mechanical data that only an AFM can provide.
Due to the FlexAFM's open optical path design structure, light can pass through the FlexAFM virtually unobstructed, and the special SureAlign™ optical design of the cantilevered arm mount SA makes it simple and easy to operate the FlexAFM in liquid test environments without the need for laser alignment. When coupled with the inverted light microscope option, the FlexAFM thus becomes the ideal device for imaging and characterising cells in a physiological environment. In this example, live Rat-2 fibroblasts were imaged.
Applications of the FlexAFM in conjunction with an inverted light microscope are certainly not limited to the life sciences. The samples below are shown, with coated glass samples being analysed by both optical microscopy and AFM. the AFM provides higher resolution data on the area under observation and explains the nature of the structures seen in the optical microscope. It can also be used for coating thickness determination.
Images of PiD cells: (left) Relative comparison image of inverted light microscopy through a FlexAFM device and inverted light microscopy option, showing live Rat-2 fibroprogenitor cells and AFM pendant arm in cell culture medium. (Right) AFM image of cells under the same device, showing details of the Rat-2 cytoskeleton, which is readily discernible across the cell membrane of these cells.AFM image size: 70 μm × 60 μm.
Relative ratio of coated glass (top) and AFM image (bottom). Observed in the optical microscope are similarly textured structures (e.g., oval interiors) corresponding to holes in the coating, as exemplified by AFM measurements. They provide a ** method for determining the thickness of the coating, which for this example is equal to 40 nm.AFM image size: 90 μm × 35 μm.
